The present invention relates to methods for the analysis of samples by ion mobility and by ion mobility combined with mass spectrometry. The apparatus and methods for sample handling and analysis described herein are enhancements of the techniques referred to in the literature relating to mass spectrometry and ion mobility spectrometry—important tools in the analysis of a wide range of chemical compounds. Specifically, ion mobility spectrometers can be used to determine the cross section of analyte ions and mass spectrometers can be used to determine their molecular mass (more precisely the ratio of molecular mass to electric charge, hereafter referred to as mass). The analysis of samples by ion mobility spectrometry and mass spectrometry comprises three main steps—formation of gas phase ions from sample material, mobility and/or mass analysis of the ions to separate the ions from one another according to ion mobility and/or mass, and detection of the ions. A variety of means and methods exist in the fields of mass and mobility spectrometry to perform each of these three functions. The particular combination of the means and methods used in a given mobility or mass spectrometer determine the characteristics of that instrument.
To mass analyze ions, for example, one might use magnetic (B) or electrostatic (E) analysis, wherein ions passing through a magnetic or electrostatic field will follow a curved path. In a magnetic field, the curvature of the path will be indicative of the momentum-to-charge ratio of the ion. In an electrostatic field, the curvature of the path will be indicative of the energy-to-charge ratio of the ion. If magnetic and electrostatic analyzers are used consecutively, then both the momentum-to-charge and energy-to-charge ratios of the ions will be known and the mass of the ion will thereby be determined. Other well known mass analyzers are the quadrupole (Q), the ion cyclotron resonance (ICR), the time-of-flight (TOF), and the Paul ion trap analyzers. More recently, linear quadrupole ion traps [J. Schwartz, M. Senko, and J. Syka, J. Am. Soc. Mass Spectrom. 13, 659 (2002); J. Hager, Rapid Commun. Mass Spectrom. 16, 512 (2002)] have become more wide spread. A new analyzer, the orbitrap, based on the Kingdon trap [K. Kingdon, Phys. Rev. 21, 408 (1923)] was recently described by A. Makarov [Q. Hu et al., J Mass Spectrom. 40, 430 (2005)]. The analyzer used in conjunction with the means and method described here may be any of a variety of these.
Before mass analysis can begin, gas phase ions must be formed from a sample material. If the sample material is sufficiently volatile, ions may be formed by electron ionization (EI) or chemical ionization (CI) of the gas phase sample molecules. Alternatively, for solid samples (e.g., semiconductors, or crystallized materials), ions can be formed by desorption and ionization of sample molecules by bombardment with high energy particles. Further, Secondary Ion Mass Spectrometry (SIMS), for example, uses keV ions to desorb and ionize sample material. In the SIMS process a large amount of energy is deposited in the analyte molecules, resulting in the fragmentation of fragile molecules. This fragmentation is undesirable in that information regarding the original composition of the sample (e.g., the molecular weight of sample molecules) will be lost.
For more labile, fragile molecules, other ionization methods now exist. The plasma desorption (PD) technique was introduced by Macfarlane et al. (R. D. Macfarlane, R. P. Skowronski, D. F. Torgerson, Biochem. Biophys. Res Commun. 60 (1974) 616)(“McFarlane”). Macfarlane discovered that the impact of high energy (MeV) ions on a surface, like SIMS would cause desorption and ionization of small analyte molecules. However, unlike SIMS, the PD process also results in the desorption of larger, more labile species (e.g., insulin and other protein molecules).
Additionally, lasers have been used in a similar manner to induce desorption of biological or other labile molecules. See, for example, Cotter et al. (R. B. VanBreeman, M. Snow, R. J. Cotter, Int. J. Mass Spectrom. Ion Phys. 49 (1983) 35; Tabet, J. C.; Cotter, R. J., Tabet, J.C., Anal. Chem. 56 (1984) 1662; or R. J. Cotter, P. Demirev, I. Lys, J. K. Olthoff, J. K.; Lys, I.: Demirev, P.: Cotter et al., R. J., Anal. Instrument. 16 (1987) 93). Cotter modified a CVC 2000 time-of-flight mass spectrometer for infrared laser desorption of non-volatile biomolecules, using a Tachisto (Needham, Mass.) model 215G pulsed carbon dioxide laser. The plasma or laser desorption and ionization of labile molecules relies on the deposition of little or no energy in the analyte molecules of interest.
The use of lasers to desorb and ionize labile molecules intact was enhanced by the introduction of matrix assisted laser desorption ionization (MALDI) (K. Tanaka, H. Waki, Y. Ido, S. Akita, Y. Yoshida, T. Yoshica, Rapid Commun. Mass Spectrom. 2 (1988) 151 and M. Karas, F. Hillenkamp, Anal. Chem. 60 (1988) 2299). In the MALDI process, an analyte is dissolved in a solid, organic matrix. Laser light of a wavelength that is absorbed by the solid matrix but not by the analyte is used to excite the sample. Thus, the matrix is excited directly by the laser, and the excited matrix sublimes into the gas phase carrying with it the analyte molecules. The analyte molecules are then ionized by proton, electron, or cation transfer from the matrix molecules to the analyte molecules. This process (i.e., MALDI) is typically used in conjunction with time-of-flight mass spectrometry (TOFMS) and can be used to measure the molecular weights of proteins in excess of 100,000 Daltons.
Atmospheric Pressure Ionization (API) includes a number of ion production means and methods. Typically, analyte ions are produced from liquid solution at atmospheric pressure. One of the more widely used methods, known as electrospray ionization (ESI), was first suggested for use with mass spectrometry by Dole et al. (M. Dole, L. L. Mack, R. L. Hines, R. C. Mobley, L. D. Ferguson, M. B. Alice, J. Chem. Phys. 49, 2240, 1968). In the electrospray technique, analyte is dissolved in a liquid solution and sprayed from a needle. The spray is induced by the application of a potential difference between the needle and a counter electrode. The spray results in the formation of fine, charged droplets of solution containing analyte molecules. In the gas phase, the solvent evaporates leaving behind charged, gas phase, analyte ions. This method allows for very large ions to be formed. Ions as large as 1 MDa have been detected by ESI in conjunction with mass spectrometry (ESMS).
In addition to ESI, many other ion production methods might be used at atmospheric or elevated pressure. For example, MALDI has recently been adapted by Laiko et al. to work at atmospheric pressure (Victor Laiko and Alma Burlingame, “Atmospheric Pressure Matrix Assisted Laser Desorption”, U.S. Pat. No. 5,965,884, and Atmospheric Pressure Matrix Assisted Laser Desorption Ionization, poster #1121, 4th International Symposium on Mass Spectrometry in the Health and Life Sciences, San Francisco, Aug. 25-29, 1998) and by Standing et al. at elevated pressures (Time of Flight Mass Spectrometry of Biomolecules with Orthogonal Injection+Collisional Cooling, poster #1272, 4th International Symposium on Mass Spectrometry in the Health and Life Sciences, San Francisco, Aug. 25-29, 1998; and Orthogonal Injection TOFMS Anal. Chem. 71(13), 452A (1999)). The benefit of adapting ion sources in this manner is that the ion optics (i.e., the electrode structure and operation) in the mass analyzer and mass spectral results obtained are largely independent of the ion production method used.
The elevated pressure MALDI source disclosed by Standing differs from what is disclosed by Laiko et al. Specifically, Laiko et al. disclose a source intended to operate at substantially atmospheric pressure. In contrast, the source disclosed by Standing et al. is intended to operate at a pressure of about 70 mtorr.
More recently, Takats et al. [Z. Takats, J. M. Wiseman, B. Gologan, and R. G. Cooks, Science 306, 471 (2004)] introduced yet another atmospheric pressure ionization method known as desorption electrospray ionization (DESI). According to Takats et al., DESI is a method for producing ions from analyte on a surface. Electrosprayed charged droplets and ion of solvent are directed at the surface under study. The impact of the charged droplets on the surface results in the desorption and ionization of the analyte to form gas phase analyte ions.
Analyte ions produced via an API method need to be transported from the ionization region through regions of differing pressures and ultimately to a mass analyzer for subsequent analysis (e.g., via time-of-flight mass spectrometry (TOFMS), Fourier transform mass spectrometry (FTMS), etc.). In some prior art sources, this was accomplished through use of a small orifice between the ionization region and the vacuum region. In other prior art, dielectric capillaries have been used to transmit ions entrained in a carrier gas from a high pressure ion production region into the vacuum chamber of mass spectrometers—see, for example, Fenn et al., U.S. Pat. No. 4,542,293 and Whitehouse et al., U.S. Pat. No. 5,844,237. In U.S. Pat. No. 6,777,672, incorporated herein by reference, Park describes a multiple section capillary for interfacing various ion production means and for transporting ions into the vacuum chamber of a mass spectrometer.
Importantly, ions are carried through the transfer capillary by entrainment in gas which is pumped from the ion production region, through the capillary, into the first vacuum region of the mass spectrometer. Typically, the gas pressure at the capillary inlet is about one atmosphere whereas the pressure at the capillary outlet, into the first pumping region, is between one and three millibar. Under these conditions, the velocity of the gas in the capillary is about 100 m/s. It is the “force” associated with this high velocity gas that is able to drive the ions away from the electrically attractive potential at the capillary entrance and towards the electrically repulsive potential at the capillary exit.
Once through the capillary, analyte ions are guided by a combination of gas flows and electric fields through differential pumping regions to a mobility and/or mass analyzer. There the ions are analyzed and detected so as to yield mobility and/or mass spectra. Any known mass or mobility analyzer or combination of mass or mobility analyzers including time-of-flight, quadrupole, Paul trap, linear ion trap, orbitrap, electric or magnetic sector, ion cyclotron resonance analyzers, drift cell, differential mobility analyzer, or trapped ion mobility analyzer might be used.
In co-pending application Ser. No. 13/152,363, incorporated herein by reference, the present inventor discloses an abridged RF quadrupole for the transport and mass analysis of ions. The abridged RF quadrupole design comprises a multitude of electrode structures arranged rectilinearly and symmetrically about a central axis (designated the z-axis). Voltages are applied to the electrode structures generating an electric field inside the abridged RF quadrupole having the form:
                              Φ          ⁡                      (                          x              ,              y              ,              t                        )                          =                                                            -                                                      Φ                    o                                    ⁡                                      (                    t                    )                                                              ·              x              ·              y                                      2              ⁢                              r                o                2                                              +                                                    E                x                            ⁡                              (                t                )                                      ·            x                    +                                                    E                y                            ⁡                              (                t                )                                      ·            y                    +          c                                    (        1        )            
where Φ is the electric field potential, Ex and Ey are functions of time relating to a homogeneous dipole field, c and ro are constants, and x and y are coordinates. Φ0 may be any function of time, t, however, as an example, is given by:
                                                        Φ              0                        ⁡                          (              t              )                                =                                                    V                ⁢                sin                            (                              2                ⁢                                                                  ⁢                π                ⁢                                                                  ⁢                ft                            )                        +            U                          ,                            (        2        )            
where V and U are RF and DC potentials respectively. Thus, in accordance with equation (1), the abridged RF quadrupole can support an RF quadrupole field which tends to focus ions to an axis—i.e. the z-axis—and a dipole field which can be used to excite ions or otherwise apply a force to the ions orthogonal to the z-axis.
The “mobility” of analyte ions through a gas can be measured under the influence of a static uniform electric field (drift type IMS). Such ion mobility spectrometers are described in detail in the literature [see, for example, G. Eiceman and Z. Karpas, Ion Mobility Spectrometry (CRC. Boca Raton, Fla. 1994); and Plasma Chromatography, edited by T. W. Carr (Plenum, New York, 1984)]. At low electric field strengths—e.g. a few kilovolts per cm—the speed of analyte ions through a gas is measured. To start the measurement, ions are pulsed into the entrance of the mobility analyzer. In the mobility analyzer, a uniform electric field accelerates the ions towards the end of the analyzer. Collisions with gas in the analyzer tend to dampen the ion motion. The action of the electric field and collision of ions with the gas thus results in an average ion speed through the gas. At the far end of the analyzer, the ions strike a detector and are detected. By measuring the time between the introduction of ions into the analyzer and the detection of the ions, the speed of the ions, and therefore their mobility, can be determined.
At low field strengths, the mobility of an ion is a constant relating the speed of the ion to the strength of the electric field. However, at high electric field strengths, the mobility of the ions varies with electric field strength. This gives rise to field asymmetric ion mobility spectrometry (FAIMS)—an extension of IMS which takes advantage of the change in ion mobility at high field strengths. FAIMS is described in detail in the literature [I. Buryakov, E. Krylov, E. Nazarov, and U. Rasulev, Int. J. Mass Spectrom. Ion Phys. 128. 143 (1993); D. Riegner, C. Harden, B. Carnahan, and S. Day, Proceedings of the 45th ASMS Conference on Mass Spectrometry and Allied Topics, Palm Springs, Calif., Jun. 1-4, 1997, p. 473; B. Carnahan, S. Day, V. Kouznetsov, M. Matyjaszczyk, and A. Tarassov, Proceedings of the 41st ISA Analysis Division Symposium, Framingham, Mass., Apr. 21-24, 1996, p. 85; and B. Carnahan and A. Tarassov, U.S. Pat. No. 5,420,424].
In recent years, IMS and FAIMS spectrometers have been combined with mass spectrometry. In U.S. Pat. No. 5,905,258 Clemmer and Reilly combine IMS with a time of flight mass spectrometer (TOFMS). This provides for a first analysis of the ions by IMS followed by a second analysis via TOFMS. Ultimately, this yields a two dimensional plot containing both the mobility and mass of the ions under investigation. The advantages of this type of combined analyzer over a mass spectrometer alone are described in detail in the literature [C. S. Srebalus et al., Anal. Chem. 71(18), 3918 (1999); J. A. Taraszka et al., J. Proteom. Res. 4, 1223 (2005); R. L. Wong, E. R. Williams, A. E. Counterman, and D. Clemmer, J. Am. Soc. Mass Spectrom. 16, 1009 (2005)] and include the separation of chemical background from analyte species for an improved signal-to-noise ratio (S/N), and the separation of ions based on compound class or charge state for easier mass spectral interpretation.
Similarly, in U.S. Pat. No. 6,504,149, for example, Guevremont et al. combine a FAIMS device with a mass spectrometer. As detailed in the literature, a combined FAIMS mass spectrometer has similar advantages as an IMS mass spectrometer [A. Shvartsburg, K. Tang, R. Smith, J. Am. Soc. Mass Spectrom. 16, 2 (2005); D. A. Barnett, B. Ells, R. Guevremont, and R. W. Purves, J. Am. Soc. Mass Spectrom. 13, 1282 (2002)]. For example, a combined FAIMS mass spectrometer has an improved signal-to-noise ratio over a mass spectrometer alone because the FAIMS device can filter away the chemical background.
Several other methods of IMS separation have been demonstrated in the prior art. For example, J. Zeleny described a parallel flow ion mobility analyzer in “J. Zeleny, Philos. Mag. 46, 120(1898).” In Zeleny's instrument, a voltage V is applied between two parallel grids separated by a distance h. Gas and ions flow through the grids parallel to the electric field. The electric field retards the motion of the ions such that the average velocity of ions between the grids is v=E·K−u, where E is the electric field strength, K is the ion mobility and u is the air flow velocity. The mobility of the ions can be calculated as K=h/Et+u/E.
In U.S. Pat. No. 5,847,386, incorporated in its entirety herein by reference, Thompson and Jolliffe suggest an ion mobility method wherein “ions are admitted into an RF multipole with an axial field, in the presence of cooling gas or drift gas, the ion velocity will reach a constant value which is proportional to the axial field. Ions of different size will drift at different velocities dependant on their shape, mass and charge, and be separated in time when they reach the exit of the device. If the exit gate . . . is opened at an appropriate time, only ions of a certain type will be admitted in the following analyzing device or other detector such as a mass spectrometer. This mobility separation may be applied to assist in the analysis of a mixture of ions . . . ”
Also recently Page et al. [J. S. Page et al., J. Mass Spectrom. 40, 1215 (2005)] and Loboda et al. [U.S. Pat. No. 6,630,662 incorporated herein in its entirety by reference] employed the parallel flow analyzer method of Zeleny in combination with RF ion optical devices. However, the method of Page et al. results in a non-uniform gas flow—i.e. the flow direction and speed is dependent on both the axial and radial position within the device. Also the DC axial electric field is non-uniform and includes radial components. Furthermore, the RF confining field generated in the Page device includes an axial component. This tends to interfere with the mobility separation and introduces a mass effect to the separation. That is, the RF field has a greater effect on ions of a given mass range and a lesser effect on ions of another. Finally, in the method of Page et al. ions of a selected mobility cannot actually be isolated from ions of greater and lesser mobility. Rather, ions of high mobility are eliminated from a stream of ions having both high and low mobility. Similarly, Loboda does not teach a means and method whereby ions from a continuous ion source can be effectively introduced into an RF device for mobility analysis.
In U.S. Pat. No. 7,838,826 B1 (M. A. Park, 2008) and in co-pending application Ser. Nos. 13/094,102, 13/094,128, and 13/094,146, ion mobility spectrometers are presented, the size of which amounts to about ten centimeters only. It is based upon moving gases driving ions over an electric counter-field barrier in a modified ion funnel built into a time-of-flight mass spectrometer. Thus, this instrument essentially roots back to J. Zeleny cited above. Unlike these many other trials to build small ion mobility spectrometers, the device by M. A. Park has already achieved ion mobility resolutions in excess of 100, and even considerably higher resolutions can be expected by future improvements.
Yet another prior art method of mobility analysis is known as differential mobility analysis (DMA). An example of such an analyzer was described by Tammet (Tammet, H. “The limits of air ion mobility resolution.” Proc. 11th Int. Conf. Atmos. Electr., NASA, MSFC, Alabama, pp 626-629 (1999).) as using the “method of inclined grids”. As described by Tammet, “The orientation of the electric field in the analyzer is . . . not perpendicular to the air flow as assumed in traditional mobility analyzers . . . . As distinct from the Zeleny grid instrument, [the analyzer of Tammet] has inlet and outlet slits for ions like traditional DMA-s. The ions to be separated do not pass through the grids and there is no harmful effect of adsorption of ions on the grids . . . . An essential advantage of the method is that the grids suppress the turbulence and maintain the plug flow profile.” Furthermore, Tammet claims the method provides improved resolution over traditional mobility analyzers.
In a similar vein, N. Agbonkonkon [N. Agbonkonkon, Counter-flow Ion Mobility Analysis Design, Instrumentation, and Characterization, Ph.D. Dissertation, Brigham Young University, December 2007] suggested the method of “counter-flow ion mobility analysis” (CIMA). According to Agbonkonkon “ . . . a high electric field (over 2500 V/cm) and high counter-gas velocity (over 10 m s-1) provide opposing forces to stop the motion of the [selected mobility] ions in space. A third force, which is stronger than either of the opposing forces is used to quickly move the ions to the detector. This third force is orthogonal to both the opposing electric field and gas flow.” In one embodiment according to Agbonkonkon, shown in FIG. 1, sample ions are passed between two perforated cylinders. The force on ions due to gas flowing from the inner cylinder to the outer cylinder is counteracted by an electric field between the cylinders.
Of the above cited prior art mobility analyzers, only DMA and CIMA are able to filter ions according to their mobility. That is, DMA and CIMA can provide a continuous ion beam of a selected mobility. However, the DMA and CIMA analyzers described in the prior art transmit ions only inefficiently because there is no mechanism to actively retain selected ions. Also, the prior art Zeleny and Agbonkonkon devices suffer from having substantially non-uniform gas velocities, resulting in poor ion mobility resolutions. Furthermore, it is not possible to operate these prior art devices in such a manner that ions are transmitted through the devices without mobility analysis—i.e. transmitting ions of all mobilities through the devices. It is therefore part of the purpose of the present invention to provide a device and method whereby ions can be mobility filtered and efficiently transmitted through the device. Another purpose of the present invention is to provide a device wherein the ions are separated according to their mobility with high resolution using guided gas flows with non-uniform gas velocity. It is a further purpose to provide a device and method whereby ions can be either mobility filtered and transmitted or efficiently transmitted without mobility filtering. Other shortcomings in prior art devices described above limit their mobility resolution, the sensitivity of the instruments employing them—i.e. many ions of interest are lost due to poor efficiency in the mobility device—and the accuracy of the mobility determinations. It is, in part, the purpose of the present invention to overcome these prior art limitations.